Isolation and amplification by polymerase chain reaction DNA of Babesia microti and Babesia divergens in ticks in Poland.

Babesia microti and B. divergens, the etiological agents of human babesiosis, are transmitted by the bite of Ixodes ricinus. The purpose of this study was differentiation of those two species in ticks collected in urban woods in the city Szczecin (north-western Poland). The prevalence of the DNA of Babesia were investigated by PCR amplification with primers to the fragment from a gene encoding the nuclear small-subunit ribosomal RNA (SS-rDNA). We examined a total of 533 specimens of Ixodes ricinus. The mean infection rate was 16.3%. Our results indicate that a B. microti and B. divergens--specific PCR test may provide a sensitive tool also for the laboratory diagnosis of human babesiosis.

The occurrence DNA of Babesia microti in ticks Ixodes ricinus in the forest areas of Szczecin.

Babesia microti, an intraerythrocytic protozoan and the etiological agent of human babesiosis, is transmitted by the bite of the tick, Ixodes ricinus. The aim of the present study was to confirm the presence of B. microti by detection of the DNA of these protozoans. The prevalence of B. microti was studied using the PCR method with primers complementary to the gene fragment encoding nuclear small-subunit ribosomal RNA (ss-rDNA). In the course of this study a total of 2095 ticks, Ixodes ricinus, were examined. The mean infection rate was 6.2%. Variable prevalance values were also obtained from six different locations and they were further modified by the seasons of the year. The results confirmed the competence of I. ricinus as a vector of B. microti and that a B. microti-specific PCR can provide a sensitive test for laboratory detection of babesiosis.

[Babesosis--difficulty of diagnosis]. / Babesjoza--trudnosci diagnostyczne.

Human babesiosis is caused predominantly by B. microti and B. divergens, a protozooan parasites of red blood cells. Both are transmitted by Ixodes ricinus ticks, also the primary vector of Lyme disease. Clinical manifestation varied widely from asymptomatic infection to a serve rapidly fatal disease. The diagnosis of babesiosis include examination of stained blood smers, serological evaluation indirect antibody tests and PCR. With the evolution PCR--based techniques, the diagnosis and monitoring of babesial infections became more sensitive and reliable.

Argyrophilic nucleolar organizer regions, bromodeoxyuridine labelling index and DNA ploidy in squamous cell carcinoma of the uterine cervix.

There is increasing evidence that rapidly proliferating tumours, i.e. those with a high bromodeoxyuridine labelling index (BrdUrdLI), could benefit from an accelerated course of radiotherapy. Also, DNA ploidy may be a prognostic factor in term of patients survival. Thus, measurements of cell kinetics and DNA ploidy might become part of routine characterization of tumours before treatment. It is supposed, that a simple and cheap argyrophilic nucleolar organizer regions (AgNOR) test reflects the proliferative status of the tumour and correlates with BrdUrdLI. The BrdUrdLI, AgNOR test and DNA ploidy were assessed in 49 squamous cell carcinoma (SCC) of the cervix (stage II B-III B) and 5 normal epithelium. The number of NORs per cell nucleus, the mean AgNOR particle area and the total AgNOR area per cell were evaluated. Significant differences in the proliferative rate were found within the examined groups of tumours assessed by the BrdUrdLI and AgNOR test. The mean BrdUrdLI values were significantly lower in normal than in carcinomatous cells, while for AgNOR values this was true for stage III B only. The mean number of AgNORs and total AgNOR area per cell were not significantly higher at stage III B than at stage II B, respectively. A high DNA aneuploidy was found in the examined tumours: 78% in stage II B and 77% in stage III B of disease. The results of proliferative markers were not significantly different in diploid than in aneuploid tumours. A significant correlation (p < 0.0001) was found between the mean AgNOR values and BrdUrdLI, however the correlation coefficient was poor (r = 0.50). This was due to different fragments of the same tumours used in these tests. Therefore these techniques might be used as independent methods reflecting the proliferative rate of the tumour.

Micronucleus assay in three transplantable mouse tumors.

The cytokinesis-block micronucleus (MN) assay was performed in three mouse tumors: two sarcomas (SaL, MCA) and Lewis lung carcinoma (LLC). To determine the optimal culture durations and cytochalasin B (cyt-B) concentrations to yield the highest proportion of binucleate cells (BC) for each tumor, the influence of the cyt-B concentration (1, 2 and 3 micrograms/ml) and culture duration (24-96) were studied. The amount of BC and the MN frequency was investigated for the different radiation doses (0-4 Gy). Dose response curves were constructed using the optimal culture duration and cyt-B concentration for each tumor. This was 24 h of incubation for MCA and 48 h for SaL and LLC and 2 micrograms/ml of cyt-B. The tumors examined differ in the mean number of spontaneously (0 Gy) occurring MN in binucleate cells. These were 0.008, 0.022 and 0.044 for MCA, SaL and LLC, respectively. The MN frequency increased with radiation dose. LLC was found to be the most radiosensitive, while MCA proved to be the least radiosensitive tumor. The average number of MN/BC at 2 Gy of irradiation (after subtraction of the value at 0 Gy) ranged from 0.05 to 0.36. The highest mean value -0.36 was shown in LLC, the middle-0.20 in SaL, and the lowest-0.05 in MCA tumor. After higher doses of irradiation numerous BC with two and more MN were found in LLC tumor, while they were not frequently observed in MCA tumor. We did not observe an increase in the MN frequency with culture duration or proliferation rate of the tumor cells. MCA has the shortest potential doubling time (Tpot) and had the lowest MN frequency from three examined tumors. The MN assay has promise to be a rapid predictive assay of radiosensitivity.